Chediak-Higashi Syndrome: a rare disorder of lysosomes and lysosome related organelles

Chediak-Higashi Syndrome (CHS) is a rare autosomal recessive disorder characterized by severe immunologic defects including recurrent bacterial infections, impaired chemotaxis and abnormal natural killer (NK) cell function. Patients with this syndrome exhibit other symptoms such as an associated lymphoproliferative syndrome, bleeding tendencies, partial albinism and peripheral neuropathies. The classic diagnostic feature of CHS is the presence of huge lysosomes and cytoplasmic granules within cells. Similar defects are found in other mammals, the most well studied being the beige mouse and Aleutian mink. A positional cloning approach resulted in the identification of the Beige gene on chromosome 13 in mice and the CHS1/LYST gene on chromosome 1 in humans. The protein encoded by this gene is 3801 amino acids and is highly conserved throughout evolution. The identification of CHS1/Beige has defined a family of genes containing a common BEACH motif. The function of these proteins in vesicular trafficking remains unknown.     

Chediak–Higashi Syndrome: A Rare Disorder of Lysosomes and Lysosome Related Organelles

Chediak–Higashi Syndrome (CHS) is a rare autosomal recessive disorder characterized by severe immunologic defects including recurrent bacterial infections, impaired chemotaxis and abnormal natural killer (NK) cell function. Patients with this syndrome exhibit other symptoms such as an associated lymphoproliferative syndrome, bleeding tendencies, partial albinism and peripheral neuropathies. The classic diagnostic feature of CHS is the presence of huge lysosomes and cytoplasmic granules within cells. Similar defects are found in other mammals, the most well studied being the beige mouse and Aleutian mink. A positional cloning approach resulted in the identification of the Beige gene on chromosome 13 in mice and the CHS1/LYST gene on chromosome 1 in humans. The protein encoded by this gene is 3801 amino acids and is highly conserved throughout evolution. The identification of CHS1/Beige has defined a family of genes containing a common BEACH motif. The function of these proteins in vesicular trafficking remains unknown.     

Reverse genetic screen for loss‐of‐function mutations uncovers a frameshifting deletion in the melanophilin gene accountable for a distinctive coat color in Belgian Blue cattle

In the course of a reverse genetic screen in the Belgian Blue cattle breed, we uncovered a 10‐bp deletion (c.87_96del) in the first coding exon of the melanophilin gene (MLPH), which introduces a premature stop codon (p.Glu32Aspfs*1) in the same exon, truncating 94% of the protein. Recessive damaging mutations in the MLPH gene are well known to cause skin, hair, coat or plumage color dilution phenotypes in numerous species, including human, mice, dog, cat, mink, rabbit, chicken and quail. Large‐scale array genotyping undertaken to identify p.Glu32Aspfs*1 homozygous mutant animals revealed a mutation frequency of 5% in the breed and allowed for the identification of 10 homozygous mutants. As expression of a colored coat requires at least one wild‐type allele at the co‐dominant Roan locus encoded by the KIT ligand gene (KITLG), homozygous mutants for p.Ala227Asp corresponding with the missense mutation were excluded. The six remaining colored calves displayed a distinctive dilution phenotype as anticipated. This new coat color was named ‘cool gray’. It is the first damaging mutation in the MLPH gene described in cattle and extends the already long list of species with diluted color due to recessive mutations in MLPH and broadens the color palette of gray in this breed.     

Drosophila mauve Mutants Reveal a Role of LYST Homologs Late in the Maturation of Phagosomes and Autophagosomes

Chediak–Higashi syndrome (CHS) is a lethal disease caused by mutations that inactivate the lysosomal trafficking regulator protein (LYST). Patients suffer from diverse symptoms including oculocutaneous albinism, recurrent infections, neutropenia and progressive neurodegeneration. These defects have been traced back to over‐sized lysosomes and lysosome‐related organelles (LROs) in different cell types. Here, we explore mutants in the Drosophila mauve gene as a new model system for CHS. The mauve gene (CG42863) encodes a large BEACH domain protein of 3535 amino acids similar to LYST. This reflects a functional homology between these proteins as mauve mutants also display enlarged LROs, such as pigment granules. This Drosophila model also replicates the enhanced susceptibility to infections and we show a defect in the cellular immune response. Early stages of phagocytosis proceed normally in mauve mutant hemocytes but, unlike in wild type, late phagosomes fuse and generate large vacuoles containing many bacteria. Autophagy is similarly affected in mauve fat bodies as starvation‐induced autophagosomes grow beyond their normal size. Together these data suggest a model in which Mauve functions to restrict homotypic fusion of different pre‐lysosomal organelles and LROs.     

Cloning of bovine LYST gene and identification of a missense mutation associated with Chediak-Higashi syndrome of cattle

An inheritable bleeding disorder with light coat color caused by an autosomal recessive gene has been reported in a population of Japanese black cattle. The disease has been diagnosed as Chediak-Higashi Syndrome (CHS) of cattle which correspond to a human inheritable disorder caused by mutation in LYST gene. To characterize the molecular lesion causing CHS in cattle, cDNAs encoding bovine LYST were isolated from a bovine brain cDNA library. The nucleotide and deduced amino acid sequences of bovine LYST had 89.6 and 90.2% identity with those of the human LYST gene, respectively. In order to identify the mutation within the LYST gene causing CHS in cattle, cDNA fragments of the LYST gene were amplified from an affected animal by RT-PCR and their nucleotide sequences were completely determined. Notably, a nucleotide substitution of A to G transition, resulting in an amino acid substitution of histidine to arginine (H2015R) was identified in the affected animal. The presence of the substitution was completely corresponding with the occurrence of the CHS phenotype among 105 members of pedigrees of the Japanese black cattle and no cattle of other populations had this substitution. These findings strongly suggested that H2015R is the causative mutation in CHS of Japanese black cattle. Received: 25 May 1999 / Accepted: 26 July 1999     

Comparing total body lipid content of young‐of‐the‐year Steller sea lions among regions of contrasting population trends

Steller sea lion (Eumetopias jubatus) young‐of‐the‐year (YOY) are nutritionally dependent upon their dam through the majority of their first year. Several indices of body condition were measured in YOY 1.5–9 mo of age captured in Southeast Alaska (n = 122), the Gulf of Alaska (n = 182), and the Aleutian Islands (n = 38) to test the hypothesis that nutritional stress impacted the ability of adult female Steller sea lions to adequately nourish their late gestation YOY in the central Aleutian Islands in the early 2000s. Body mass (kg) and percent total body lipid content (%TBL) increased with age in all three regions of Alaska that were sampled (P < 0.05). Young‐of‐the‐year 7–9 mo of age were leaner in Southeast Alaska (27.6% ± 1.0%) and Gulf of Alaska (29.5% ± 0.8%) than in the Aleutian Islands (35.7% ± 1.2%, P < 0.001). Condition indices calculated from morphometric measures did not strongly predict the %TBL measured by isotope dilution. The trend for Aleutian Island YOY to have larger body mass and larger body fat reserves are counter to what would be expected if dams were unable to adequately provision their late lactation YOY due to inadequate food availability in the central Aleutian Islands.     

Interspecific crossing between blue‐tailed damselflies Ischnura elegans and I. senegalensis in the laboratory

Ischnura species (Odonata) are among the most common damselflies in the world, which often exhibit female color polymorphisms. One morph, called androchrome, is similar to males in its color pattern, whereas the other morphs, generally referred to as gynochromes, exhibit female‐specific colors. In several Ischnura species, the female polymorphism is heritable, although molecular and genetic mechanisms remain largely unknown. The dominant‐recessive patterns of the female color morphs may differ between species. For example, androchromic females are dominant to gynochromic females in Ischnura elegans, whereas androchromic females are recessive in Ischnura senegalensis. Here we report a case of interspecific hybridization between a gynochrome female of I. elegans and a male of I. senegalensis in the laboratory. We obtained 61 hybrid adult offspring, of which all 31 females were of gynochrome morph. DNA analyses of the hybrids confirmed that nuclear DNA sequences were derived from both parent species, whereas mitochondrial DNA sequences were maternally inherited. In the hybrids, the postocular spots of female heads, the shape of male appendages, and the color of female's cerci resembled those of I. elegans, whereas the size of abdominal blue spots was similar to that of I. senegalensis. The shape of prothorax and basal abdominal markings were intermediate in females. The larval developmental traits and the morphological changes in the final larval instar of the hybrids were similar to those of I. senegalensis. To our knowledge, this is the first report of hybrids between two damselfly species with different dominant‐recessive patterns of female color morphs.     

A TIR–NBS protein encoded by Arabidopsis Chilling Sensitive 1 (CHS1) limits chloroplast damage and cell death at low temperature

Survival of plants at low temperature depends on mechanisms for limiting physiological damage and maintaining growth. We mapped the chs1‐1 (chilling sensitive1‐1) mutation in Arabidopsis accession Columbia to the TIR‐NBS gene At1g17610. In chs1‐1, a single amino acid exchange at the CHS1 N‐terminus close to the conserved TIR domain creates a stable mutant protein that fails to protect leaves against chilling stress. The sequence of another TIR‐NBS gene (At5g40090) named CHL1 (CHS1‐like 1) is related to that of CHS1. Over‐expression of CHS1 or CHL1 alleviates chilling damage and enhances plant growth at moderate (24°C) and chilling (13°C) temperatures, suggesting a role for both proteins in growth homeostasis. chs1‐1 mutants show induced salicylic acid production and defense gene expression at 13°C, indicative of autoimmunity. Genetic analysis of chs1‐1 in combination with defense pathway mutants shows that chs1‐1 chilling sensitivity requires the TIR‐NBS‐LRR and basal resistance regulators encoded by EDS1 and PAD4 but not salicylic acid. By following the timing of metabolic, physiological and chloroplast ultrastructural changes in chs1‐1 leaves during chilling, we have established that alterations in photosynthetic complexes and thylakoid membrane integrity precede leaf cell death measured by ion leakage. At 24°C, the chs1‐1 mutant appears normal but produces a massive necrotic response to virulent Pseudomonas syringae pv. tomato infection, although this does not affect bacterial proliferation. Our results suggest that CHS1 acts at an intersection between temperature sensing and biotic stress pathway activation to maintain plant performance over a range of conditions.     

Interaction of MC1R and PMEL alleles on solid coat colors in Highland cattle

Six solid colors occur in Highland cattle: black, dun, silver dun and red, yellow, and white. These six coat colors are explained by a non‐epistatic interaction of the genotypes at the MC1R and PMEL genes. A three base pair deletion in the PMEL gene leading to the deletion of a leucine from the signal peptide is observed in dilute‐colored Highland cattle (c.50_52delTTC, p.Leu18del). The mutant PMEL allele acts in a semi‐dominant manner. Dun Galloway cattle also have one copy of the deletion allele, and silver dun Galloway cattle have two copies. The presence of two adjacent leucine residues at the site of this deletion is highly conserved in human, horse, mouse and chicken as well as in cattle with undiluted coat colors. Highland and Galloway cattle thus exhibit a similar dose‐dependent dilution effect based on the number of PMEL :c.50_51delTTC alleles, as Charolais cattle with PMEL :c.64G>A alleles. The PMEL :c.64G>A allele was not found in Highland or Galloway cattle.     

Cloning,characterization and localization of <Emphasis Type="Italic">CHS</Emphasis> gene from blood orange, <Emphasis Type="Italic">Citrus sinensis</Emphasis> (L.) Osbeck cv. Ruby

Chalcone synthase (CHS) is involved in the biosynthesis of anthocyanin. In this study, a full-length DNA of CHS gene (named as CsCHS-bo) was cloned from the blood orange, Citrus sinensis (L.) Osbeck cv. Ruby. The gene was 1,512 bp in size containing an open reading frame (1,176 bp) encoding 391 amino acids. Comparative and bioinformatic analyses revealed that the deduced protein of CsCHS-bo was highly homologous to CHS from other plant species. The protein of CsCHS-bo had four CHS-specific conserved motifs and a CHS-family signature sequence GFGPG. Phylogenetic analysis indicated that the protein of CsCHS-bo was in a subgroup with CHS of Ruta Palmatum. The CsCHS-bo was localized to the chromosomes 2p, 4p and 6p by an improved fluorescence in situ hybridization technique, indicating that at least three copies of CsCHS-bo were present in the genome. The novel nucleotide sequence data published here have been deposited in the EMBL/DDBJ/GenBank databases under accession number EU410483.     

Antibodies to Pseudogymnoascus destructans are not sufficient for protection against white‐nose syndrome

White‐nose syndrome (WNS) is a fungal disease caused by Pseudogymnoascus destructans (Pd) that affects bats during hibernation. Although millions of bats have died from WNS in North America, mass mortality has not been observed among European bats infected by the fungus, leading to the suggestion that bats in Europe are immune. We tested the hypothesis that an antibody‐mediated immune response can provide protection against WNS by quantifying antibodies reactive to Pd in blood samples from seven species of free‐ranging bats in North America and two free‐ranging species in Europe. We also quantified antibodies in blood samples from little brown myotis (Myotis lucifugus) that were part of a captive colony that we injected with live Pd spores mixed with adjuvant, as well as individuals surviving a captive Pd infection trial. Seroprevalence of antibodies against Pd, as well as antibody titers, was greater among little brown myotis than among four other species of cave‐hibernating bats in North America, including species with markedly lower WNS mortality rates. Among little brown myotis, the greatest titers occurred in populations occupying regions with longer histories of WNS, where bats lacked secondary symptoms of WNS. We detected antibodies cross‐reactive with Pd among little brown myotis naïve to the fungus. We observed high titers among captive little brown myotis injected with Pd. We did not detect antibodies against Pd in Pd‐infected European bats during winter, and titers during the active season were lower than among little brown myotis. These results show that antibody‐mediated immunity cannot explain survival of European bats infected with Pd and that little brown myotis respond differently to Pd than species with higher WNS survival rates. Although it appears that some species of bats in North America may be developing resistance to WNS, an antibody‐mediated immune response does not provide an explanation for these remnant populations.     

Blood parasite infection differentially relates to carotenoid‐based plumage and bill color in the American goldfinch

Male and female American goldfinches (Spinus tristis) express condition‐dependent carotenoid‐based plumage and bill coloration. Plumage color is relatively static, as pigments incorporated into feathers during the spring molt cannot be mobilized thereafter. In contrast, bill color is dynamic, reflecting changes in condition over short time periods. Previous studies have shown that male and female ornaments, though similar in expression, are differentially related to measures of immunocompetence, suggesting that the relationship between ornamentation and parasite infection may differ between the sexes. In this study, we evaluate the relationship between condition‐dependent ornamentation (plumage and bill color) and blood parasite infection in male and female American goldfinches. We captured goldfinches after completion of the pre‐alternate molt and prior to the onset of nesting and assessed prevalence of Trypanosoma parasites via blood smears. Plumage color strongly predicted trypanosome infection: Birds with more colorful plumage were less likely to present infections. In contrast, we detected no relationship between infection and bill color, which in other studies has been shown to dynamically reflect current condition. Sex did not affect the relationship between infection status and either ornament. Together, these results suggest that physiological pathways linking carotenoid ornamentation and infection may vary even within a single species.     

The striking polyphyly of Suiriri: Convergent evolution and social mimicry in two cryptic Neotropical birds

Two species of Suiriri (Aves: Tyrannidae) inhabit semi‐open habitats in South America: the polytypic Suiriri Flycatcher (S. suiriri) and the monotypic Chapada Flycatcher (S. affinis). The phylogenetic relationship between these congeneric species has never been investigated in detail. Here we used molecular tools—three nuclear introns and two mitochondrial genes—to investigate the systematic position of the Chapada Flycatcher, comparing the results found with morphological and behavioral data. We found that the polytypic Suiriri Flycatcher to be monophyletic and that it is included in a clade of Elaeniini flycatchers including Phyllomyias, Phaeomyias, and Capsiempis among other genera. The Chapada Flycatcher, on the other hand, is a member of the Fluvicolini, sister to Sublegatus, and should be allocated on its own monospecific genus, which we herein describe. We suggest that social mimicry is responsible for the remarkable convergence in size, shape, plumage coloration, and behavior in the adults of the Suiriri Flycatcher and the Chapada Flycatcher.     

Howler monkey foraging ecology suggests convergent evolution of routine trichromacy as an adaptation for folivory

Primates possess remarkably variable color vision, and the ecological and social factors shaping this variation remain heavily debated. Here, we test whether central tenants of the folivory hypothesis of routine trichromacy hold for the foraging ecology of howler monkeys. Howler monkeys (genus Alouatta) and paleotropical primates (Parvorder: Catarrhini) have independently acquired routine trichromacy through fixation of distinct mid‐ to long‐wavelength‐sensitive (M/LWS) opsin genes on the X‐chromosome. The presence of routine trichromacy in howlers, while other diurnal neotropical monkeys (Platyrrhini) possess polymorphic trichromacy, is poorly understood. A selective force proposed to explain the evolution of routine trichromacy in catarrhines—reliance on young, red leaves—has received scant attention in howlers, a gap we fill in this study. We recorded diet, sequenced M/LWS opsin genes in four social groups of Alouatta palliata, and conducted colorimetric analysis of leaves consumed in Sector Santa Rosa, Costa Rica. For a majority of food species, including Ficus trees, an important resource year‐round, young leaves were more chromatically conspicuous from mature leaves to trichromatic than to hypothetical dichromatic phenotypes. We found that 18% of opsin genes were MWS/LWS hybrids; when combined with previous research, the incidence of hybrid M/LWS opsins in this species is 13%. In visual models of food discrimination ability, the hybrid trichromatic phenotype performed slightly poorer than normal trichromacy, but substantially better than dichromacy. Our results provide support for the folivory hypothesis of routine trichromacy. Similar ecological pressures, that is, the search for young, reddish leaves, may have driven the independent evolution of routine trichromacy in primates on separate continents. We discuss our results in the context of balancing selection acting on New World monkey opsin genes and hypothesize that howlers experience stronger selection against dichromatic phenotypes than other sympatric species, which rely more heavily on cryptic foods.     

Sex determination in the wild: a field application of loop‐mediated isothermal amplification successfully determines sex across three raptor species

PCR‐based methods are the most common technique for sex determination of birds. Although these methods are fast, easy and accurate, they still require special facilities that preclude their application outdoors. Consequently, there is a time lag between sampling and obtaining results that impedes researchers to take decisions in situ and in real time considering individuals’ sex. We present an outdoor technique for sex determination of birds based on the amplification of the duplicated sex‐chromosome‐specific gene Chromo‐Helicase‐DNA binding protein using a loop‐mediated isothermal amplification (LAMP). We tested our method on Griffon Vulture (Gyps fulvus), Egyptian Vulture (Neophron percnopterus) and Black Kite (Milvus migrans) (family Accipitridae). We introduce the first fieldwork procedure for sex determination of animals in the wild, successfully applied to raptor species of three different subfamilies using the same specific LAMP primers. This molecular technique can be deployed directly in sampling areas because it only needs a voltage inverter to adapt a thermo‐block to a car lighter and results can be obtained by the unaided eye based on colour change within the reaction tubes. Primers and reagents are prepared in advance to facilitate their storage at room temperature. We provide detailed guidelines how to implement this procedure, which is simpler (no electrophoresis required), cheaper and faster (results in c. 90 min) than PCR‐based laboratory methods. Our successful cross‐species application across three different raptor subfamilies posits our set of markers as a promising tool for molecular sexing of other raptor families and our field protocol extensible to all bird species.     

Contrasting the seasonal and elevational prevalence of generalist avian haemosporidia in co‐occurring host species

Understanding the ecology and evolution of parasites is contingent on identifying the selection pressures they face across their infection landscape. Such a task is made challenging by the fact that these pressures will likely vary across time and space, as a result of seasonal and geographical differences in host susceptibility or transmission opportunities. Avian haemosporidian blood parasites are capable of infecting multiple co‐occurring hosts within their ranges, yet whether their distribution across time and space varies similarly in their different host species remains unclear. Here, we applied a new PCR method to detect avian haemosporidia (genera Haemoproteus, Leucocytozoon, and Plasmodium) and to determine parasite prevalence in two closely related and co‐occurring host species, blue tits (Cyanistes caeruleus, N = 529) and great tits (Parus major, N = 443). Our samples were collected between autumn and spring, along an elevational gradient in the French Pyrenees and over a three‐year period. Most parasites were found to infect both host species, and while these generalist parasites displayed similar elevational patterns of prevalence in the two host species, this was not always the case for seasonal prevalence patterns. For example, Leucocytozoon group A parasites showed inverse seasonal prevalence when comparing between the two host species, being highest in winter and spring in blue tits but higher in autumn in great tits. While Plasmodium relictum prevalence was overall lower in spring relative to winter or autumn in both species, spring prevalence was also lower in blue tits than in great tits. Together, these results reveal how generalist parasites can exhibit host‐specific epidemiology, which is likely to complicate predictions of host–parasite co‐evolution.     

Species‐dependent microarchitectural traits of iridescent scales in the triad taxa of Ornithoptera birdwing butterflies

Ornithoptera birdwing butterflies have blue, green, or orange iridescent scales in different species or subspecies. To understand the species‐ or subspecies‐dependent scale color differences, we performed comparative morphometric analyses of iridescent scales from three closely related taxa: O. priamus priamus (green), O. priamus urvillianus (blue), and O. croesus (orange). The three types of Ornithoptera wings exhibited reversible color changes to longer wavelengths with different kinetics upon immersion in methanol, suggesting that their color differences are at least partly based on differences in the size of air cavities made by nanostructures. Cover scales of all three color types were visually semi‐transparent glass scales that exhibited color when placed on a dark background. The dorsoventral differences in coloration were observed in single scales, suggesting the optical importance of scale surfaces. Scanning electron microscopy of cover scales in cross section revealed that all color types exhibited finely sculpted tapered ridges and thick, irregular basal multilayers containing tandemly clustered granular objects and air cavities. Scale thickness, ridge height, and multilayer thickness were significantly different among the three color types, and granular object size was significantly different between orange scales and blue and green scales. We conclude that each of the three taxa of Ornithoptera butterflies possesses unique quantitative size values on tapered ridges and irregular multilayers with granular objects and air cavities to express unique structural color. These species‐ or subspecies‐dependent structural colors might have evolved via quantitative shifts in these microarchitectural traits rather than via changes in the basic developmental or architectural plan for color expression.     

A genome scan of diversifying selection in Ophiocordyceps zombie‐ant fungi suggests a role for enterotoxins in co‐evolution and host specificity

Identification of the genes underlying adaptation sheds light on the biological functions targeted by natural selection. Searches for footprints of positive selection, in the form of rapid amino acid substitutions, and the identification of species‐specific genes have proved to be powerful approaches to identifying the genes involved in host specialization in plant‐pathogenic fungi. We used an evolutionary comparative genomic approach to identify genes underlying host adaptation in the ant‐infecting genus Ophiocordyceps, which manipulates ant behaviour. A comparison of the predicted genes in the genomes of species from three species complexes—O. unilateralis, O. australis and O. subramanianii—revealed an enrichment in pathogenesis‐associated functions, including heat‐labile enterotoxins, among species‐specific genes. Furthermore, these genes were overrepresented among those displaying significant footprints of positive selection. Other categories of genes suspected to be important for virulence and pathogenicity in entomopathogenic fungi (e.g., chitinases, lipases, proteases, core secondary metabolism genes) were much less represented, although a few candidate genes were found to evolve under positive selection. An analysis including orthologs from other entomopathogenic fungi in a broader context showed that positive selection on enterotoxins was specific to the ant‐infecting genus Ophiocordyceps. Together with previous studies reporting the overexpression of an enterotoxin during behavioural manipulation in diseased ants, our findings suggest that heat‐labile enterotoxins are important effectors in host adaptation and co‐evolution in the Ophiocordyceps entomopathogenic fungi.     

Phenotypic and genetic diversity in aposematic Malagasy poison frogs (genus Mantella)

Intraspecific color variation has long fascinated evolutionary biologists. In species with bright warning coloration, phenotypic diversity is particularly compelling because many factors, including natural and sexual selection, contribute to intraspecific variation. To better understand the causes of dramatic phenotypic variation in Malagasy poison frogs, we quantified genetic structure and color and pattern variation across three closely related species, Mantella aurantiaca, Mantella crocea, and Mantella milotympanum. Although our restriction site‐associated DNA (RAD) sequencing approach identified clear genetic clusters, they do not align with current species designations, which has important conservation implications for these imperiled frogs. Moreover, our results suggest that levels of intraspecific color variation within this group have been overestimated, while species diversity has been underestimated. Within major genetic clusters, we observed distinct patterns of variation including: populations that are phenotypically similar yet genetically distinct, populations where phenotypic and genetic breaks coincide, and populations that are genetically similar but have high levels of within‐population phenotypic variation. We also detected admixture between two of the major genetic clusters. Our study suggests that several mechanisms—including hybridization, selection, and drift—are contributing to phenotypic diversity. Ultimately, our work underscores the need for a reevaluation of how polymorphic and polytypic populations and species are classified, especially in aposematic organisms.